Description (applicant's description): Our long-term goal is to determine the atomic structures of herpes simplex virus type 1 (HSV-1) particles at different maturational stages and to elucidate their interactions with cellular factors. An intact HSV-1 virion is spherical with a diameter of about 2000 A and consists of several compartments: glycoproteins, membrane, tegument layer, capsid and dsDNA. Because of its size and complexity, we plan to employ a combination of electron cryomicroscopic, x-ray crystallographic, computational, biochemical, and genetic approaches to study the entire virion, the capsid and the individual capsid proteins. The capsid has a diameter of 1250 A and has a T=16 icosahedral lattice symmetry. The capsid shell has a total mass of 0.2 billion daltons and is made up of four different proteins (VP5, VP19C, VP23 and VP26) with molecular masses of 149, 50, 34 and 12 kDa respectively. This proposal will be carried out in collaboration with Frazer Rixon, a molecular virologist and Florante Quiocho, a crystallographer and will utilise established resources in the form of intact and mutant virions and different types of naturally occurring and recombinant capsid particles. We have six specific aims. The first is to obtain the crystal structures of individual molecular components of the capsid shell and to dock them into the intermediate resolution (6-8 A) capsid structure determined by electron cryomicroscopy. The second is to extend the structural study of the entire B capsid shell towards 6 A resolution by electron cryomicroscopy. The third is to integrate data from a variety of crystallographic, computational and labeling experiments to interpret the 6 A capsid map in terms of secondary structure elements, specific residue locations and possible folds. The fourth is to follow the process of capsid maturation by determining the structures of a number of intermediates from procapsid to mature virion. The fifth is to determine the identity of the icosahedrally-bound tegument protein that we have recently observed and also to characterize its structure at a higher resolution than at present. The sixth is to explore the possibilities for studying the intact capsid at atomic resolution by crystallography and electron cryomicroscopy. This will be the largest icosahedral particle to be studied in such detail. The outcome of our investigations will provide insights on the structural assembly of this important human pathogen and also advance the structural approaches for studying large macromolecular machines.